Analysis of chromosome conformation capture combined with high-throughput sequencing (4C-seq)

 

w4CSeq is a computational analysis pipeline for robust characterization of the physical organization around selected promoters and other functional elements using both enzyme-based and sonication-based 4C-Seq.

Circular chromosome conformation capture, when coupled with next-generation sequencing (4C-Seq), can be used to identify genome-wide interaction of a given locus (a “bait” sequence) with all of its interacting partners. Conventional 4C approaches used restriction enzyme digestion to fragment chromatin, and recently sonication approach was also applied for this purpose. However, bioinformatics pipelines for analyzing sonication-based 4C-Seq data are not well developed. In addition, data consistency as well as similarity between the two methods has not been explored previously. 

From biological replicates, we found good correlation (r>0.6) for inter-chromosomal interactions identified in either enzyme or sonication method. Compared to enzyme approach, sonication method generated less distal intra-chromosomal interactions, possibly due to the difference in chromatin fragmentation. From all mapped interactions, we further applied statistical models to identify enriched interacting regions. Interestingly, data generated from the two methods showed 30% overlap of the reproducible interacting regions. The interacting sites in the reproducible regions from both methods are similarly enriched with active histone marks.

Both enzyme-based and sonication-based 4C-Seq methods are valuable tools to explore long-range chromosomal interactions. Due to the nature of sonication-based method, correlation analysis of the 4C interactions with transcription factor binding should be more straightforward.

w4CSeq applies a customized pipeline to deal with both enzyme digestion and sonication fragmentation based 4C-seq data, identifies statistically significant regions, draws interacting plots for intra-chromosome and inter-chromosome interactions, and overlays genomic features (TSS, TTS, CpG sites), DNA replication timing and user-provided annotation onto it. The combined plot will help uncover significant features in/around 4C regions.

 

Availability

w4CSeq is currently being tested internally, and is not available for public use yet.

 

Reference

  • Gao F, Zong W, Lu W, Wang K. "Comparative analysis of 4C-Seq data generated from enzyme-based and sonication-based methods." BMC Genomics 2013, 14:345
  • Cai M, Gao F, Lu W, Wang K. "W4CSEQ -- a server to process 4C-Seq data." (In preparation)